Tissue shrinkage and stereological studies

Shrinkage often takes place in biological tissues during the different phases of preparation for microscopy. This can have detrimental effects on the stereological estimates, even when unbiased procedures are used. There are different types of shrinkage, and an awareness of them is essential when designing stereological studies. Some forms of shrinkage can be taken into account to ensure the unbiasedness of an estimator, but some cannot and should be avoided. Dimensional changes that take place during fixation, embedding, sectioning, mounting, and staining can seriously compromise one's ability to make assumption-free estimates of length and surface, but there are steps that can be taken to reduce the impact of these changes on estimates of object number and size. This article describes types of shrinkage and the effects of shrinkage on estimators of object number. It gives an example of how to make a number-weighted correction of section thickness and also discusses the consequences of shrinkage for the validity of estimates of object size.